1a. Objectives (from AD-416):
1. Determine the extent to which dietary antioxidants alter obesity-induced and/or exercise-induced changes in mitochondrial function and insulin sensitivity. Sub-objective 1A. Determine the influence of anti-oxidant supplementation on changes in insulin sensitivity induced in the rat by high dietary fat and exercise. Sub-objective 1B. Determine the degree to which anti-oxidant supplementation alters exercise-induced changes in insulin sensitivity and mitochondrial function responses of overweight/obese individuals. 2. Identify sites and causes of obesity-induced and exercise-induced oxidative stress. Sub-objective 2A. Determine the effects of obesity and exercise on the temporal and cellular activation of the nuclear factor (erythroid-derived 2)-like 2 (Nrf-2)/Anti-oxidant Response Element pathway. Sub-objective 2B. Identify and characterize obesity-induced and exercise-induced oxidative changes to insulin signaling pathway proteins. 3. Identify, characterize and compare sites of obesity-induced versus exercise-induced mitochondrial respiratory changes. Sub-objective 3A. Determine the degree to which anti-oxidant supplementation blunts exercised-induced and obesity-induced changes in mitochondria.
1b. Approach (from AD-416):
In order to complete the objectives of this proposal, we will utilize a combination of studies in humans, rodents that examine physiologic, metabolomic, genetic, and proteomic endpoints. In Objective 1, we will perform studies in humans and rodents to determine how antioxidant (vitamin E and vitamin C) supplementation affects insulin responses to exercise and obesity. The study in humans will involve analysis of exercise adaptation and insulin responses in previously untrained individuals and if antioxidant supplementation either enhances or negates these adaptations. Rodent studies will further examine molecular mechanisms underlying these adaptations. In Objective 2, we will determine the extent to which obesity, exercise, and anti-oxidant supplementation alter redox balance in animals and specific cells and to identify specific proteins whose thiol redox status is altered in obesity, exercise, and anti-oxidant supplementation. These studies will utilize transgenic mouse models and proteomic approaches. In Objective 3, we will determine the extent to which obesity, exercise, and anti-oxidant supplementation alter mitochondrial function. These studies will utilize rat models of exercise and obesity. Whole tissue and isolated mitochondria will be studied for changes in total mitochondrial content, mitochondrial gene expression, and respiration, and mitochondrial enzyme activities.
3. Progress Report:
During FY 2012, the project team made progress in multiple areas. Objective 1A. We performed studies in rats testing the hypothesis that vitamin C and vitamin E supplementation prevents glucose intolerance in obese rats. We are currently analyzing the samples and data. We will test the hypothesis that vitamin C and vitamin E supplementation prevents exercise adaptations in rats in the FY 2013. This work will examine the role of anti-oxidant supplements as adjuncts to treatment of obesity or enhancing the effects of exercise. Objective 1B. This study investigates whether anti-oxidant supplementation (vitamins C and E) modulate exercise responses particularly with respect to blood sugar regulation in overweight/obese people. Unfortunately, the scientist overseeing this study left the GFHNRC. Over 6 months of recruiting, we had poor response. It was decided to halt recruitment and the study. Objective 2A. This objective tests the hypothesis that exercise and nutritional factors regulate cellular redox status in tissue specific ways. To test this hypothesis, we developed a colony of Anti-oxidant Reporter Element (ARE)-reporter mice. We were able to see induction of this pathway in liver through feeding the mice butylated hydroxyanisole (BHA), a known ARE inducer. However, whether with forced or voluntary exercise, we were unable to demonstrate changes in ARE pathways in liver or skeletal muscle. Owing to the poor results with this mouse model, we decided not to pursue these experiments further in this model. Demonstrated that adipocyte differentiation reduces protein levels of the ARE-dependent protein NQO1. We demonstrated that the dietary component sulforaphane blocks adipocytes hypertrophy in part through induction of NQO1. This work is currently in press - Free Radical Biology and Medicine. Objective 2B. The hypothesis of this work is that lack of exercise and obesity induces a hyper-reductive state of insulin signaling proteins. We developed methods for determining changes in protein redox state and demonstrated that glutathione modification of proteins was decreased in visceral adipose and liver of obese rats. This work was accepted for publication in the journal Obesity. Objective 3. We performed exercise studies in mice (voluntary) and rats (forced running wheel). Analysis of mitochondria DNA and mitochondrial respiratory proteins in skeletal muscle did not demonstrate a difference between sedentary and exercised animals. This research project includes an ancillary project examining the role of Atlantic salmon consumption as a means to increase omega 3 fatty acids in humans. This project is a collaboration with the ARS-National Cold Water Aquaculture Center in Franklin, ME. We published one paper demonstrating that baking salmon decreases the presence of fatty acid oxidation products and does not decrease the content of beneficial omega-3 fatty acids in salmon – Journal of Agricultural and Food Chemistry. We have submitted one paper demonstrating that eating baked salmon increases blood omega-3’s in a dose-dependent manner.
1. Obesity regulates protein modification in visceral adipose and liver. This work studied how obesity alters oxidative damage to proteins. This work focused on protein oxidation in liver, muscle, and visceral adipose – tissues that contribute to insulin resistance in obesity. Data generated by ARS scientists at Grand Forks, ND demonstrate that a specific type of protein modification – glutathionylation-is actually reduced by induction of obesity. These results indicate that protein changes in obesity are more complex than originally thought and provide insight into how oxidative stress and protein function are modulated in obesity. Because of the study the effects of anti-oxidant supplements upon protein damage in obesity will be investigated.
2. Fat cell growth and antioxidants. During fat cell growth levels of anti-oxidant proteins are reduced, but the protein levels can be maintained by the dietary component sulforophane. ARS scientists at Grand Forks, ND determined that dietary sulforaphane blocks adipocyte development in part by inducing the production of NQO1, an anti-oxidant protein. The results provide insight into how oxidative stress is modulated in obesity and that how diet components may reduce obesity.
3. Farmed salmon consumption increases plasma levels of n3 fatty acids in humans. Two servings of oily fish per week are recommended to prevent heart disease by increasing levels of n3 fatty acids in humans. However there were no studies showing the effect of eating this level of fish upon n3 fatty acids in people. ARS scientists at Grand Forks, ND showed that consumption of 4 ozs of farmed Atlantic salmon twice per week significantly increased n3 fatty acids. This work has impact for consumers, health professionals, and for producers of farmed salmon.Raatz, S.K., Young, L.R., Picklo, M.J., Sauter, E.R., Qin, W., Kurzer, M.S. 2012. Total dietary fat and fatty acid content modify plasma phospholipid fatty acids, desaturase activity indices and urinary prostaglandin E. Nutrition Research. 32:1-7.